Targeting the Exon2 splice cis-element in PD-1 and its effects on lymphocyte function

Abstract

T-cell therapies have proven to be a promising treatment option for cancer patients in recent years, especially in the case of chimeric antigen receptor (CAR)-T cell therapy. However, the therapy is associated with insufficient activation of T cells or poor persistence in the patient’s body, which leads to incomplete elimination of cancer cells, recurrence, and genotoxicity. By extracting the splice element of PD-1 pre-mRNA using biology based on CRISPR/dCas13 in this study, our ultimate goal is to overcome the above-mentioned challenges in the future. PD-1 plays an important role in controlling T cell responses and is expressed at the cell surface of T cells following activation. The receptor PD-1 interferes with T cell receptor (TCR) signaling following interaction with PD-L1. The outcome of stimulation via PD-1 leads to decreases in cytokine secretion and cell proliferation. We extracted the RNA region of PD-1 pre-mRNA using CD8+T cell lines and examined the effect of targeting the Exon2 splice cis-element on the production of cytokines in the present study. In particular, the production of IFN-γ, TNF-α, GM-CSF was lower in RNA-targeted cells than in non-targeted cells, but the cytokine secretion capacity and cell proliferation were maintained in RNA-targeted cells. These results suggested that the use of the RNA editing technology, CRISPR/dCas13 strategy offers a novel approach to mitigate genotoxicity in lymphocytes with cytokine production and cell proliferation.